Overview
Hydrogels are an important platform technology for biomedical solutions in many areas, including managing chronic ailments, such as rheumatoid arthritis and osteoporosis, and treating acute conditions, such as hemorrhaging and cancer. Their utility is due in part to their high water content and tunable mechanical properties, which makes them inherently similar to living tissue. Hydrogels have been widely investigated for contact lenses, as scaffolds for load-bearing connective tissue, and as materials for controlled drug release. Their demonstrated mechanical behavior spans a broad spectrum of useful properties: chemically crosslinked double-network hydrogels exhibit high toughness, and physically crosslinked protein-based hydrogels can shear-thin and are well suited for injectable biomaterials. This latter property is particularly promising in the clinical setting because the ability to implant a solid formulation using a minimally-invasive technique can enable superior control in the delivery of drugs and cellular materials.
A variety of injectable hydrogel formulations have been developed, such as those that rely on responsive chemical crosslinking or physical association triggered in the physiological environment. Shear-thinning gels are particularly useful due to their ability to achieve full elastic strength rapidly after injection and promote the survival of encapsulated cells due to plug flow profiles during injection. Unfortunately, current shear-thinning gels do not meet the requirements for elastic modulus and fracture toughness required for load-bearing applications, and there is no method to prevent shear-thinning after injection. To maintain injectability during processing and ultimately achieve the mechanical properties desired at the implantation site, a responsive toughening mechanism for shear-thinning hydrogels is needed.
Protein-Based Physical Hydrogels as Injectable Biomaterials
Genetic engineering and biosynthesis provide a route to the facile design and production of polymers with precisely determined sequences, and these tools have been leveraged in the engineering of a number of useful materials for sensing, responsive drug delivery, and catalysis. A variety of naturally occurring protein sequences have been utilized in physical hydrogels, including coiled-coil associating domains and elastin-based polypeptides exhibiting lower critical solution temperatures. In the case of proteins incorporating coiled-coil domains, precise sequence control allows gels to be prepared with enhanced control over network topology. Depending on their amino acid sequence, these alpha-helical domains will associate in either parallel or anti-parallel directions and do so with well-defined valencies. When linked together by flexible chains, either structure-less polyelectrolyte protein domains or poly(ethylene glycol), they reversibly associate to form a network. Thus, preparing hydrogels with different coiled-coils or different sized linker domains provides a handle on the material's mechanical properties and degradation rates.
A well-studied coiled-coil-based gel is composed of helices that associate in pentameric bundles (abbreviated P) connected by 10 repeat units of a structure-less nonapeptide linker sequence (abbreviated C). The linker sequences can be modified to include cell binding peptides without sacrificing the mechanical properties of the gel, providing a useful mechanism for engineering biofunctionality. PC10P gels prepared at a 7% (w/v) concentration have elastic moduli near 4 kPa, typical for physically associating gels. The material is responsive to pH and thermal changes that cause the protein to unfold, but these responsive transitions are typically outside of physiological ranges. At large strain the gels exhibit a yield response indicative of rapid shear-thinning behavior, and recovery of the elastic modulus is almost instantaneous following cessation of shear. Velocity profiles observed in capillary flow have plug flow character due to shear banding near the boundaries. Because of this localization of strain, hydrogel formulations encapsulating cells sustain high survival rates post-injection. Similar success in cell survivability has been achieved in other peptide-based hydrogels, although in this example the recovery of the elastic modulus was on the order of minutes.
Gel Toughening
One method for the toughening of hydrogels is the incorporation of dispersed domains of stiff material, as in clay or carbon nanotube (CNT) nanocomposites. These materials have tremendous promise to enhance mechanical properties—such as elastic modulus—at low loadings, and they are compatible with typical polymer processing techniques. In the case of CNT nanocomposites, the slow progress in achieving the desired enhancement has been due to the difficulty of producing molecularly disperse composites. For CNTs and other materials used in nanocomposites, aggregated clusters can lead to lower surface areas and higher flexibility of the filler material, reducing the expected reinforcement effect. Although surface functionalization has been used to attempt to resolve this issue, care must be taken such that the chemical processing does not compromise the morphology of the filler.
Double network hydrogels represent an alternative means to achieve toughened gels, and they have had great success approaching the elastic modulus and fracture properties of articular cartilage. These gels consist of two independent, interpenetrating networks typically of different chemical nature and connected over different length scales. Such materials can exhibit an order of magnitude improvement in elastic moduli up to tens of MPa and stresses at fracture up to 20 times that of a single network of either one of the component polymers. A large mismatch in monomer feed ratios and crosslinking densities is important to achieve optimal enhancement. Optical interference measurements on compression studies show that optimized double-network gels exhibit weak birefringence compared to sub-optimal formulations. These data suggest that stresses are effectively distributed throughout the material, with the loosely crosslinked network dissipating localized stresses and preventing crack growth in the material.
While the above strategies provide valuable lessons on engineering toughness in soft materials, neither has been suitably adapted for use in injectable biomaterials, which require toughening mechanisms that can be activated after implantation, when flow behavior is no longer needed. High performance clay nanocomposites and double-network hydrogels have been prepared from chemical gels, so responsive crosslinking of a liquid formulation has been proposed to meet these constraints. However, responsive chemical gels typically have higher cytotoxicity and slower curing times than shear-thinning physical gels.
Two approaches have been taken to responsively toughen physical hydrogels. Recently, a protein-based physical hydrogel has been strengthened via photo-initiated chemical crosslinking to give approximately a three-fold improvement in the elastic modulus. However, even the crosslinked material is insufficient for use in load-bearing tissues. Additionally, photoirradiation of an implanted material can be complicated in the clinical setting due to limited tissue penetration of the radiation, and the toxicity of residual crosslinking agents must be taken into account. The modulus of a triblock-copolymer hydrogel was increased by over two orders of magnitude by crosslinking the polyanionic midblock with divalent inorganic cations. While this work demonstrates that the double-network principle can be implemented using two physical networks, crosslinking with diffusible ions is unsuitable for many biomedical applications. The material would equilibrate with the local tissue environment, likely losing some of the enhancement in mechanical performance. Furthermore, the processing required to achieve enhanced toughness is not readily adapted to the clinical setting, and the response time is 12-72 hours.
Much progress has been made in the toughening of soft materials, but no single strategy has succeeded in simultaneously satisfying the demands of injectable biomaterials, namely pressure-driven flow through narrow geometries, rapid restoration of solid-like behavior after the cessation of shear, responsive toughening, and biocompatibility. Nevertheless, lessons from the design of double-network chemical gels—the interpenetration of a stiff but brittle network with a soft but ductile network—can inform the design of responsively tough, injectable biomaterials.